RU2525181C1 - Method for preparing biodegradable membranes for preventing adhesion formation following cardiosurgical operations - Google Patents

Abstract

FIELD: food industry.

SUBSTANCE: what is described is a polymeric composition formulation of poly(hydroxy butyrate-co-hydroxyvalerate) (PHB-co-PHV) with additionally administered poly(D,L-lactide) in a solid ratio of 3:1 and dissolved in chloroform to the concentration of 6-9%; the composition is thereafter mixed for 2 hours and heated to 35°C. What is described is a membrane prepared by electrostatic formation (electrospinning) which involves including biologically active substances of fibronolytic preparations or direct action anticoagulants in the structure of fibres.

EFFECT: membranes possess biocompatible properties, a biodegradation life no more than 60 days, and enable preventing the adhesion formation effectively in experiment.

2 cl, 2 tbl

Description

The invention relates to medicine, namely to cardiovascular surgery, and can be used to prevent the development of adhesions after open heart surgery.

The formation of postoperative adhesions is a frequent complication in cardiovascular surgery, which can lead to significant complications in repeated heart operations. Prevention of the formation of postoperative adhesions is one of the necessary conditions for solving this problem, since the adhesion process lengthens the operation time, increases the frequency of postoperative complications, and causes massive bleeding. At the same time, hospital mortality in case of resternotomy reaches 14-15%, and in urgent cases 43%, fatal hemorrhage occurs in 2-6% of cases. To prevent the formation of adhesions, agents are used that differ in the mechanism of action, method of application and effectiveness. The most optimal for cardiac surgery is the use of membranes, which serve as a temporary barrier separating the wound surfaces between the pericardium and sternum.

The use of membranes based on biodegradable natural polymers makes it possible to effectively separate wound surfaces before healing, followed by biodegradation of the membrane with the formation of non-toxic products.

Known anti-adhesive membranes made using carboxymethyl cellulose and sodium hyaluronate "Seprafilm" (A novel adhesion barrier facilitates reoperations in complex congenital cardiac surgery / Walther T, Rastan A // The Journal of Thoracic and Cardiovascular Surgery. 2005. Vol.129, Is. 2.- P.359-363).

The use of such membranes reduces the incidence of adhesions after surgery in more than 50% of patients. The disadvantages of the known membranes include low efficiency in the presence of blood, which always accompanies surgery on the heart and large blood vessels.

In addition, there is information in the literature about the inflammatory reaction with Seprafilm films (Seprafilm-induced peritoneal inflammation: a previously unknown complication / Klingler PJ, Floch NR, Seelig MH et al. / Report of a case // Dis Colon Rectum - 1999. - V.42. - N.12. - P.1639-1643).

Known biodegradable membrane Repel-CV, which includes polylactic acid and polyethylene glycol (Andrew JL et al. A Novel bioresorbable film reduces postoperative adhesions after infant cardiac surgery // The Annals Thorac Surg, 2008; V.86 (2): P. 614-621). These polymers are widely used in implantable and bioresorbable medical devices. However, the results of a randomized clinical trial on the use of Repel-CV showed that in 21% of patients the membrane was insolvent due to the formation of adhesions, cases of mediastinitis were noted. In addition, during hydrolysis of the polymer chain of polylactic acid in vivo, lactic acid is released, accompanied by significant acidification of the tissues (pH shift to 3.2-3.4) and an increase in the inflammatory response of tissues.

Known anti-adhesive membrane made of a polymer of bacterial origin of the class of polyhydroxyalkanoates - poly-4-hydroxybutyrate (PHB), dissolved in 1,4 dioxane or tetrahydrofuran (Patent US No. 7943683, IPC C08G 63/06, B29C 47/00, publ. 17.05 .2011). High biocompatibility of polyoxyalkanoates is based on the fact that 3-hydroxybutyric acid is a natural metabolite of cells and tissues of animals and humans. Due to its high biocompatibility, PHB is used as a raw material for the production of absorbable sutures, osteoprostheses, surgical plates, and anti-adhesive membranes.

The disadvantages of membranes based on the PHB monopolymer include insufficient elasticity and excessive fragility, which prevents the optimal location of the membrane in the surgical wound and may contribute to the failure of the sutures during fixation of the membrane. In addition, the monopolymer solvents used are highly toxic substances and when mixed with oxygen form an explosive mixture.

Closest to the claimed technical solution is a membrane made from a polymer composition of 3-hydroxybutyrate and 3-hydroxyvalerate (3-PHB / 3-PGV), dissolved in chloroform with the addition of antibiotics or drugs from the group of non-steroidal anti-inflammatory drugs (Patent RU No. 2447902, IPC A61L 31/08, A61L 31/10, A61L 31/16). The use of a copolymer of polyhydroxybutyrate-hydroxyvalerate can increase the elasticity of the membranes, compared with the use of a monopolymer - polyhydroxybutyrate. The use of chloroform as a solvent minimizes the toxic effect of the solvent. Membranes are made by irrigation.

The disadvantages of these membranes include a long biodegradation period of more than three months. While for the prevention of adhesions, this time interval should not exceed 60 days, because according to the phases of adhesiogenesis, the formation of adhesions between injured surfaces ends by 30 days after surgery. A longer placement of the membrane in the area of surgical intervention is undesirable in view of the fact that it can cause a protective reaction of the body, as to a foreign body.

The technical result of the invention is the creation of a biologically active biodegradable membrane to prevent adhesions in patients after cardiac surgery, with increased hemocompatibility and biocompatibility, improved physicochemical properties, as well as optimal biodegradation periods in the presence of blood.

The technical result is achieved by the fact that poly (D, L-lactide) is added to the composition of the polymer composition based on the copolymer of polyhydroxybutyrate / hydroxyvalerate (PHBV), which helps to reduce the biodegradation time. In addition, a method of manufacturing a membrane using the method of electrostatic formation (electrospinning) allows you to get a membrane with a microfiber structure, which also reduces the period of biodegradation of polymers. In addition, during the electrostatic formation of the membrane, the structure of the fibers includes biologically active substances from the group of fibrinolytic drugs or drugs from the group of direct anticoagulants, thus, as the membrane is biodegradable, the release of biologically active substances occurs uniformly, and thus a prolonged local drug effect occurs.

A method for the manufacture of anti-adhesive biodegradable membranes to prevent adhesions after cardiosurgical operations, comprising dissolving a polyhydroxybutyrate / hydroxyvalerate copolymer (PHBV) in a solvent followed by the addition of biologically active substances.

The difference is that poly (D, L-lactide) is also added to the composition of the biopolymer composition in a ratio of 3: 1.

The difference is that the membrane contains biologically active substances from the group of fibrinolytic drugs or direct anticoagulants, consisting of fibrinolysin, streptokinase, streptodekase and alteplase, or from the group of direct-acting anticoagulants, consisting of unfractionated heparin, sodium enoxaparin, dalteparin and naltroparin.

The difference is that the membrane is made by electrostatic molding, which allows you to create a membrane in the form of a non-woven fabric, consisting of microfibers which contain a biologically active substance. The application of the electrostatic molding method gives the membrane a micro-dimensional structure that undergoes biodegradation faster than film membranes.

The method of electrostatic molding allows you to place a biologically active substance inside the polymer microfiber, which is released during the destruction of the membrane and has a local therapeutic prolonged effect. Since the formation of adhesions is a consequence of a decrease in the fibrinolytic activity of the body, or a consequence of the deposition of fibrin in the surgical wound, the use of biologically active substances from the group of fibrinolytic drugs will have a local fibrinolytic effect. The use of direct-acting anticoagulants will prevent the formation of fibrin and additionally have an anti-inflammatory effect, which in general will prevent the formation of adhesions. Thus, a polymer membrane made of biodegradable polymers (PHBV + poly (D, L-lactide)) will mechanically separate injured surfaces after surgery for a period of not more than 60 days, and the biologically active substance released as the membrane degrades will provide additional therapeutic directed action.

As a result of comparative studies, it was proved that the strength of samples - films made of PHBV + poly (D, L-lactide) by electrospinning is close to the strength of the native pericardium, to which the membrane is fixed during surgery, and the elasticity of membranes made by electrospinning is 20% higher than membranes made by irrigation. The results of the results are presented in table 1.

Table 1 Physico-mechanical characteristics of membranes Membrane Composition Membrane thickness mm Strength (MPa) Relative extension (%) Irrigation-made PHBV 0.1 19.31 ± 4.85 501.5 ± 58.78 PHBV made by electrospinning 0.05 6.79 ± 3.54 612.7 ± 42.64 PHBV + poly (D, L-lactide) made by electrospinning 0.05 16.25 ± 5.64 604.98 ± 29.79

The hemocompatibility of the obtained membranes was judged by the amount of hemolysis induced by the aqueous extract from the polymer membranes. It was found that membranes based on the PHBV copolymer, both made by irrigation and by electrospinning, do not adversely affect red blood cells; hemolysis was not detected in any sample.

Using scanning electron microscopy, the structure of the PHBV + poly (D, L-lactide) membrane was evaluated. The electrospinning membranes are composed of randomly spaced 3.2-3.6 microns fibers. The inclusion of biologically active substances in the composition helps to reduce the thickness of the fiber to 1.7-1.9 microns.

The tissue reaction to the membranes was studied by subcutaneous implantation of biopolymer samples in laboratory rats. After implantation, a thin connective tissue capsule is formed around the membrane, which is consistent with published data (Gogolewski et al., 1993, Chaput et al., 1995, Qu et al., 2006). All samples are characterized by the absence of signs of an inflammatory process. An in vivo study of the biodegradation of experimental films showed that membranes made by irrigation completely degrade only by the end of the sixth month after subcutaneous implantation in laboratory rats. At the same time, electrospinning membranes completely degrade 60 days after implantation without signs of lymphocytic infiltration. The inclusion of poly (D, L-lactide) in the membranes made it possible to reduce biodegradation time by 30% - only empty chambers were visualized 45 days after implantation, no membrane residues were detected.

The effectiveness of the developed membranes was evaluated under conditions of the development of adhesions in laboratory animals. Adhesive disease was simulated in male Wistar rats weighing 250-300 g, since rats from all laboratory animals have maximum fibrinolytic activity. 28 days after the operation, the presence and severity of the formed adhesions were evaluated (see table 2).

As can be seen from table 2, membranes made by electrospinning effectively prevent the formation of adhesions in 65% of cases, and only 5% of the animals observed extensive, dense adhesions. When biologically active substances are included in the membranes, it contributes to a more effective prevention of adhesion formation - in 85% of rats adhesions were not found, in 15% of animals weak adhesions were observed.

table 2 Anti-adhesion efficiency of membranes depending on manufacturing technology Membrane manufacturing technology Number of operated animals Number of adhesions % Including extensive tight adhesions % Irrigation membrane twenty 10 fifty 6 thirty Electrospinning membrane twenty 7 35 one 5 Electrospinning membrane + biologically active substances twenty 3 fifteen 0 0

The following is an example implementation of the proposed method of manufacturing a biodegradable membrane.

A portion of the powder of a copolymer of 3-PHB / 3-PGV and poly (D, L-lactide) in the ratio (3: 1) is dissolved in chloroform to a concentration of 6-9% and thoroughly mixed for 2 hours on a magnetic stirrer heated to 30-35 ° C. The resulting polymer solution is placed in a syringe dispenser, which is installed in the installation for electrostatic molding. A biologically active substance is placed in a second syringe dispenser. The fiber is formed using a coaxial nozzle, which allows you to form a polymer fiber, into which the biologically active substance is contained. The process of forming the fiber occurs at a voltage of 18-23 kV, the feed rate of the polymer and biologically active substance from 0.4 ml / h to 1 ml / h. The size of the membranes is from 13 × 13 cm to 15 × 18 cm, and the thickness is from 150 to 500 microns. Films are sterilized with ethylene oxide at room temperature.

Thus, biodegradable membranes based on a copolymer of polyhydroxybutyrate / hydroxyvalerate and poly (D, L-lactide), made by electrostatic molding, with biologically active substances from the group of fibrinolytic drugs or direct anticoagulants included in the structure of microfibers, have satisfactory biocompatible biodegradability not exceeding 60 days and can effectively prevent the formation of adhesions in the experiment.

Claims (2)

1. A method of manufacturing biodegradable membranes to prevent the formation of adhesions after cardiosurgical operations, including dissolving in chloroform a copolymer of polyhydroxybutyrate / hydroxyvalerate (PHBV) and including at least one biologically active substance in the composition, followed by stirring for 2 hours magnetic stirrer and heated to 35 ° C, characterized in that the composition of the polymer composition is additionally introduced poly (D, L-lactide), and the mixture is dissolved in chloroform to the end centration of 6–9%, while the dry solids ratio of the PHBV copolymer and poly (D, L-lactide) in the composition is 3: 1, and the membrane is made by electrostatic formation; 2. A method of manufacturing biodegradable membranes according to claim 1, characterized in that as a biologically active substance selected drugs from the group of fibrinolic drugs or direct-acting anticoagulants.